Adelostemma gracillimum (Asclepiadaceae) is an herb found in Southwest of China and Burma. Extracts from the plant have been used for thousands of years in folk medicine for therapeutic purposes. The roots of the herb are used as a nourishing and roborant tonic and in treating convulsions in children. Extracts of Adelostemma gracillimum have been found to contain pregnane glycosides (Mu et al., 1992; Gao et al., 2009).
The central nervous system (CNS) is regulated by many complex pathways which monitor crucial cellular events such as proliferation, differentiation, and apoptosis (or cell death). While apoptosis is an integral part of neuronal development, defects in the mechanisms of apoptosis are thought to contribute to the disease pathology of stroke, epilepsy, and a host of neurodegenerative diseases (Raff et al., 1993). Thus, one area of drug development is to understand the processes that underlie neuronal survival and apoptosis.
Studies have shown that many, growth factors and neurotrophins, including insulin, insulin-like growth factor-1, brain-derived neurotrophic factor (BDNF), nerve-growth factor (NGF), and neurotropins 3 and 4/5, can promote neuronal survival by mediating specific signaling cascades such as the ERK and the PI 3-kinase pathways (Segal and Greenberg, 1996). Thus, it is desirable to identify and/or to develop compounds that can promote neuronal survival against apoptosis induced by nutrient withdrawal.
NMDA receptors are ligand-gated ion channels located primarily within the CNS. They belong to the family of ionotropic glutamate receptors and exist as multiple subtypes due to the different combinations of subunits—NR1, NR2 (NR2A, NR2B, NR2C, NR2D) and NR3—that can be expressed. In addition to the agonist binding site, NMDA receptors have multiple distinct binding sites for various compounds that enhance, modulate and inhibit the activation of the receptors.
It is known that NMDA receptors are involved in neuronal communication and play important roles in synaptic plasticity and mechanisms that underlie learning and memory. Under normal conditions, NMDA receptors engage in synaptic transmission via the neurotransmitter glutamate, which regulates and refines synaptic growth and plasticity. However, when there are abnormally high levels of glutamate (i.e. under pathological conditions), NMDA receptors become over-activated, resulting in an excess of Ca2+ influx into neuronal cells, which in turn causes excitotoxicity and the activation of several signaling pathways that trigger neuronal apoptosis. Glutamate-induced apoptosis in brain tissue also accompanies oxidative stress resulting in loss of ATP, loss of mitochondrial membrane potential, and the release of reactive oxygen species and reactive nitrogen species (e.g. H2O2, NO, OONO−, O2−) causing associated cell damage and death. Decreased nerve cell function and neuronal cell death eventually occur. Excitotoxicity also occurs if the cell's energy metabolism is compromised.
Over-activation of the NMDA receptors is implicated in neurodegenerative diseases and other neuro-related conditions as it causes neuronal loss and cognitive impairment, and also plays a part in the final common pathway leading to neuronal injury in a variety of neurodegenerative disorders such as amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease and Huntington's disease, as well as conditions such as stroke. NMDA receptors are also implicated in many other neurological disorders, such as multiple sclerosis, cerebral palsy (periventricular leukomalacia), and spinal cord injury, as well as in chronic and severe mood disorders (Mathew SJ et al., Rev Bras Psiquiatr, 27:243-248 (2005)).
NMDA receptors have played crucial roles in both regulating and promoting normal nervous system functions as well as causing cell death, which leads to lethal conditions. There has been increasing evidence to show that the type of signal given to a cell depends on the location of the activated NMDA receptor. Growth and survival-promoting signals result from activated synaptic NMDA receptors, while cell death causing signals result from extrasynaptic NMDA receptors. Recent studies also indicate that activated synaptic NMDA receptors lead to robust phosphorylation of the transcription factor CREB on the transcriptional regulatory residue Ser133 and promote CREB-dependent gene expression and neuronal survival. However, activated extrasynaptic NMDA receptors transiently phosphorylate CREB and do not activate CREB-dependent gene expression, resulting in neuronal cell death (Hardingham et al., 2002).
Yet, there are few effective therapeutic agents for excitotoxicity to alleviate symptoms of its associated neuronal disorders. One complication for the development of effective NMDA antagonists as neurotherapeutic drugs is that many NMDA antagonists also exhibit psychotogenic and neurotoxic properties. For example, MK-801 (dizocilpine maleate) is capable of providing certain degree of neuroprotection in ischemic stroke, but is associated with psychotropic and adverse motor effects. Thus, it is desirable to identify and/or to develop compounds that can potentiate NMDA synaptic activity resulting in neuroprotection.
Amyloid beta (Aβ) is a cleavage product derived from the amyloid precursor protein (APP), which accumulates as extracellular or senile plaques, the characteristic hallmark of the neurodegenerative disease Alzheimer's disease (AD). While the actual cause of AD remains elusive, Aβ has been implicated in many reports to play a part in the initiation and progression of the disease (Hock et al., 2003). Furthermore, studies have shown that Aβ is neurotoxic (Hartman et al., 2005), resulting in neuronal loss and subsequent memory loss and cognitive impairment. Upon the addition of Aβ to primary neuronal cultures, apoptosis is triggered which leads to cell death (Estus et al., 1997). Caspases are a family of cysteine-aspartic acid proteases that are involved in cell apoptosis through sequential activation by proteolytic processing of inactive proenzymes to form the active enzyme. Caspase-3 is considered to be the executioner of the apoptotic pathway and is the predominant caspase involved in the cleavage of the amyloid precursor protein as well as in the production of Aβ, which is associated with neuronal death in Alzheimer's disease. Exogenous addition of Aβ into neuronal cultures initiates caspase-3 dependent apoptosis. Therefore, developing inhibitors against caspase-3 activation is one therapeutic approach in AD treatment.
Dendritic spines, or spines, are small membranous protrusions from a neuron's dendrite that typically receive input from a single synapse of an axon. Dendritic spines serve as a storage site for synaptic strength and help transmit electrical signals to the neuron's cell body. Spines, however, require maturation after formation. Immature spines have impaired signaling capabilities, and typically only have necks and lack, or have very small, “heads”. Matured spines maintain both heads and necks. Spines with strong synaptic contacts typically have a large spine head, which connect to the dendrite via a membranous neck (mushroom shape) (Yuste and Denk, 1995). Decreased spine density has been reported in aged neurons of the CA1 area of hippocampus as well as the layer III pyramidal layer (Duan et al., 2003; von Bohlen und Halbach et al., 2006). In stressed animals, an overall shift in the population of spines is observed with a reduction in large spines and an increase in small spines in the prefrontal cortex (Radley et al., 2008). Spine reduction is also associated with major depression as well as in schizophrenia (Law et al., 2004). Cognitive disorders such as autism, mental retardation, Fragile X Syndrome, stroke, and chronic alcoholism may be resultant from abnormalities in dendritic spines, especially in regards to the number of spines and their maturity (Bhatt et al., 2009; von Bohlen und Halbach et al., 2009).
Epilepsy is a clinical phenomenon consisting of excessive neuronal activities in the brain which are manifested by seizures (Fisher et al., 2005). Epileptic seizures can take on the form of tonic or clonic movements accompanied sporadically by convulsions and other neurological and physical and psychic symptoms. Seizures occur transiently and can occur through provocation, though not necessarily (Aylwar R., 2008). The incidence of epilepsy is estimated at approximately 0.3 to 0.5 percent in different populations throughout the world, with the prevalence of epilepsy estimated at 5 to 10 people per 1000 which makes it one of the most prevalent neurological disorders (Shinnar and Pellock, 2002).
Epilepsy is classified by etiology, observable seizure activity, location of seizure activity in the brain, accompanying medical symptoms, and the initial event which provoked the seizure activity. The main characteristic that distinguishes the different categories of seizures is whether the seizure activity is partial (synonymous with focal) or generalized (Brodie et al., 2009). Currently, there are over 40 recognized epilepsy types classified by type of seizures, EEG recordings, physical manifestations, treatment and prognosis (Badawy et al., 2009). In vivo experiments have shown that genetic mutations in both voltage and receptor-gated ion channels are responsible for various types of seizure activities (Meisler M, and Keamey J., 2005). For example, dysregulation in voltage-gated sodium channel localized in GABAergic neurons have been implicated in severe myoclonic seizures of infants (SMEI) (Yu et al., 2009). Seizures can also occur from trauma such as ischemia or hypoxia, fever, and encephalitis, which then lead to dysregulation of balance in neuronal transmission (Bialer and White, 2010). Neuroinflammation has also been linked to the pathophysiology of epilepsy where inflammatory responses caused by cytokines lead to neuronal damage and changes in the neuronal environment, resulting in the hyperactivity observed in seizures (Ravizza et al., 2008).
Drugs to treat epilepsy are based on anticonvulsant medications. Currently, there are more than 20 antiepileptic drugs available. However, they have been reported to have side effects on patients including mood changes, sleepiness, or unsteadiness in gait (Schmitz, 2006). While the new generation of antiepileptic drugs has considerable improvements in terms of safety, tolerability and pharmacokinetics to control epileptic seizures, an estimated 30% of patients suffer from pharmacoresistant epilepsy and thus fail to respond to multiple medications (Andres and Antoaneta, 2007). The development of new drug candidates is necessary to offer alternative targets for the control of seizures with fewer or no side effects, and better efficacy is required to offer complete control of seizures in epileptic patients.
Therefore, there is a need to identify and/or to develop compounds that are capable of (i) preventing and/or treating CNS disorders, such as excitotoxicity, epilepsy, neurodegenerative diseases and neuropathological conditions; (ii) promoting neuronal survival against apoptosis induced by nutrient withdrawal; and (iii) enhancing the brain's cognitive functions. The present invention satisfies this and other needs.